Summary of Research for the entire reporting period (in terms of CPR questions)
The overall goal of this project was to determine the effect of radiation, especially cosmic radiation, on atherosclerosis, the disease that results in heart attacks and strokes. This could help to estimate the risk for astronauts on deep-space missions, such as a trip to Mars. This project consisted of two arms. The cell component, using cultured human aortic endothelial cells (HAECs) was designed to elucidate the mechanism by which absorption of radiation, either photons or ions, causes pro-atherogenic changes in the vascular endothelium. The animal component, which used development of atherosclerotic plaques as an endpoint, was to determine the eventual consequences of these changes for development of cardiovascular disease, and to work toward identification of early endpoints that predict later consequences.
ApoE -/- mice were used in the animal component of this project because they already had a well-characterized response to x-ray radiation, especially with respect to atherosclerosis. Previous studies had established that this mouse model reproduces the pathology seen in humans after radiation exposure. Moreover, these mice mimic an adult human population in that they unavoidably develop some degree of asymptomatic atherosclerotic disease spontaneously with age.
In this project, we made progress on three CPR questions, as detailed below.
Degen-1, “How can tissue specific risk models be developed for the major degenerative tissue risks, including heart, circulatory, endocrine, digestive, lens and other tissue systems in order to estimate GCR (galactic cosmic radiation) and SPE (solar particle event) risks for degenerative diseases?”
The tissue of interest for this project is the vascular system, especially the major vessels of the chest and neck, which are the major sites of clinically significant atherosclerosis.
To address Degen-1, we established that 56Fe (i.e., iron ions, a particularly damaging component of cosmic radiation) targeted to the aorta and carotid arteries has similar effects as x-rays, but at a lower dose. That is, while 8-14 Gy x-rays have been shown to exacerbate atherosclerosis in this mouse model, 56Fe has significant effects at as little as 2 Gy. The fact that 56Fe can produce effects at lower doses is not surprising, since particle radiation can be particularly damaging. In radiobiology, the relative biological effect (RBE) is defined as the ratio of absorbed dose of a radiation in question to the absorbed dose of a reference radiation (usually gamma- or x-rays) required to produce the same biological effect in a particular tissue. Based on this definition, the RBE of approximately 4-7 for 56Fe ion-induced atherosclerosis (signifying that iron ion causes similar adverse cardiovascular effects at a 4-7 fold lower dose than x-rays) is consistent with those associated with other effects.
It is known from human epidemiologic data that x-ray or gamma-ray doses as low as 1 Gy can increase the risk of cardiovascular disease, suggesting that humans are 8-14 times more sensitive to the pro-atherogenic effects of high-energy photon radiation as apo-E -/- mice. Assuming an RBE of 4-7 for humans as well as mice, the prediction would be that 56Fe as low as 0.14-0.25 Gy might significantly increase the risk for astronauts. Further work will be needed to confirm that this extrapolation from mice to humans is valid. Although 56Fe targeted to the aortic arch accelerated development of atherosclerosis, the dose dependence of 56Fe-induced atherosclerosis varied by site. While 5 Gy was required for a significant effect in the aortic arch, 2 Gy was sufficient in the aortic root (where the aorta emerges from the heart) and carotid arteries. These differences might be due to pro-atherogenic contributions of local factors, such as shear stress (the force felt by the inside wall of the vessel from blood flowing by). This advances our understanding of the how radiation induces atherosclerosis by demonstrating that, while direct radiation effects on the vessel are clearly a component of the 56Fe effect on atherosclerosis, the risk can be modified by other local factors.
The above experiments were done by irradiating only the heart and major blood vessels of mice. Since the entire astronaut will be irradiated during spaceflight, we then asked the question of whether radiation-induced atherosclerosis is primarily dependent on local effects on the vessels themselves or whether significant contributions, either positive or negative, from other organ systems might be important when the whole body is exposed. At least a component of the pro-atherogenic effect of 56Fe was a direct effect on the vessel itself, because radiation did not increase atherosclerosis in un-irradiated areas of the aorta. However, potential effects on the immune system may either exacerbate or ameliorate the local radiation effects. Therefore, we compared whole-body irradiation to that targeted to the chest and neck. Our data indicate that, as with direct, local factors, the influence of systemic factors also depends on the site and perhaps on the particular effect measured. Thickening of the carotid artery wall, a hallmark of atherosclerosis, was largely unaffected by 56Fe irradiation of the rest of the body. Plaque formation in the aortic arch, however, seemed to be slightly blunted in whole-body 56Fe irradiated mice vs. those with radiation targeted to the major vessels. This was a minor effect however, because significant exacerbation of plaques was seen at 2 Gy in either case. We conclude that the major effect of radiation is on the vessels themselves, but that systemic effects from irradiation of other tissues may modify the damage to a small extent.
Degen-2, “What are the mechanisms of degenerative tissues risks in the heart, circulatory, endocrine, digestive, lens, and other tissue systems? What surrogate endpoints do they suggest?” The cell biology arm of this study (Aim #2) is specifically designed to elucidate the mechanism of radiation-induced atherosclerosis. It is known that gamma- and x-radiation increase the adhesiveness of vascular endothelium, a key, early step in the pathogenesis of atherosclerosis. We established that 56Fe increases adhesiveness as well. It is hypothesized that this increased endothelial adhesiveness initiates endovascular inflammation in atherosclerosis-prone arteries, and that this inflammatory response can become self-perpetuating. Although a common mechanism for making cells stick to each other is to increase the number of adhesion molecules on the surface of the cell, we showed that x-ray-induced endothelial adhesiveness can occur even without up-regulation of adhesion molecules. We further showed that the mechanism by which this occurs is dependent upon chemokine signaling. We determined that the integrin alpha4beta1 (also known as VLA-4), an important adhesion molecule needed for atherosclerosis to develop, is one of the adhesion molecules activated by radiation. This was not a direct effect, but was mediated by production of signaling molecules, known as chemokines, by cells in the blood vessels.
Experiments at Brookhaven National Laboratory (where particle accelerators can mimic components of cosmic radiation) then confimed that, despite the major differences in how x-rays and heavy ions interact with tissue, 56Fe also increases endothelial adhesiveness through a chemokine-dependent mechanism. Moreover, the 56Fe radiation-induced adhesiveness also depends on signaling to the integrin alpha4beta1.
Degen-3, “What are the progression rates and latency periods for degenerative risks, and how do progression rates depend on age, gender, radiation type, or other physiological or environmental factors?”
Degen-3 is addressed by Aims #1 and #3. Since Aim #1 involves experiments with both x-rays and 56Fe, progression rates for these two radiation types can be compared. We showed earlier that x-rays and 56Fe have similar pro-atherogenic effects, increasing adhesiveness of human aortic endothelial cells. The dose of 56Fe required to do this was much lower than that required for x-rays, however.
We also demonstrated a linear energy transfer (LET) dependence of radiation-induced atherosclerotic changes in the major blood vessels. Thus, to estimate risk for astronauts on deep space missions it might be better to consider not only the total dose of heavy ion likely to be absorbed, but to take into account the mix of ions and their energies as well.